CN111647505B - DNA acquisition swab and preparation method and application thereof - Google Patents
DNA acquisition swab and preparation method and application thereof Download PDFInfo
- Publication number
- CN111647505B CN111647505B CN202010537018.0A CN202010537018A CN111647505B CN 111647505 B CN111647505 B CN 111647505B CN 202010537018 A CN202010537018 A CN 202010537018A CN 111647505 B CN111647505 B CN 111647505B
- Authority
- CN
- China
- Prior art keywords
- swab
- dna
- polycaprolactone
- hydrophilic medium
- solution
- Prior art date
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- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 28
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Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M33/00—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus
- C12M33/02—Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus by impregnation, e.g. using swabs or loops
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J9/00—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof
- C08J9/28—Working-up of macromolecular substances to porous or cellular articles or materials; After-treatment thereof by elimination of a liquid phase from a macromolecular composition or article, e.g. drying of coagulum
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2201/00—Foams characterised by the foaming process
- C08J2201/04—Foams characterised by the foaming process characterised by the elimination of a liquid or solid component, e.g. precipitation, leaching out, evaporation
- C08J2201/048—Elimination of a frozen liquid phase
- C08J2201/0482—Elimination of a frozen liquid phase the liquid phase being organic
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2305/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2301/00 or C08J2303/00
- C08J2305/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2367/00—Characterised by the use of polyesters obtained by reactions forming a carboxylic ester link in the main chain; Derivatives of such polymers
- C08J2367/04—Polyesters derived from hydroxy carboxylic acids, e.g. lactones
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2401/00—Characterised by the use of cellulose, modified cellulose or cellulose derivatives
- C08J2401/08—Cellulose derivatives
- C08J2401/26—Cellulose ethers
- C08J2401/28—Alkyl ethers
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2405/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2401/00 or C08J2403/00
- C08J2405/04—Alginic acid; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2405/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2401/00 or C08J2403/00
- C08J2405/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2467/00—Characterised by the use of polyesters obtained by reactions forming a carboxylic ester link in the main chain; Derivatives of such polymers
- C08J2467/04—Polyesters derived from hydroxy carboxylic acids, e.g. lactones
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2479/00—Characterised by the use of macromolecular compounds obtained by reactions forming in the main chain of the macromolecule a linkage containing nitrogen with or without oxygen, or carbon only, not provided for in groups C08J2461/00 - C08J2477/00
- C08J2479/02—Polyamines
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Abstract
The invention provides a DNA acquisition swab, a preparation method and application thereof. The preparation raw materials of the DNA acquisition swab comprise polycaprolactone, a hydrophilic medium and a carrier; the hydrophilic medium comprises a saccharide and/or a non-saccharide organic high molecular polymer. The DNA collecting swab takes polycaprolactone as a framework, and a saccharide and/or a non-saccharide organic high polymer is taken as a hydrophilic medium to modify the polycaprolactone framework, and is compounded on a carrier together to form the swab, so that the formed swab has excellent adsorption and collection effects on cells and trace DNA, can release more DNA in the subsequent DNA extraction step, does not influence the subsequent PCR amplification, and can be applied to detection of contact trace DNA.
Description
Technical Field
The invention relates to the field of biomedical materials, in particular to a DNA acquisition swab, a preparation method and application thereof.
Background
Forensic DNA analysis is a scientific technology for analyzing the distribution and transmission rule of DNA genetic markers in a population by using a modern DNA technology, determining the consistency and genetic relationship of analysis samples, and providing evidence for detecting broken cases and judicial judgment. The autosomal STR (short tandem repeat) locus is the most widely used genetic marker in the current paternity test laboratory, and a plurality of commercial STR detection kits for forensic DNA analysis have been developed at present, so that scientific application of investigation and case breaking and judicial judgment is greatly promoted. However, in the process of performing inspection by applying a large amount of these technologies, there is a problem that the existing inspection conditions cannot be well solved, such as the detection of trace, degraded and mixed inspection materials, such as ropes, shells, masks, mixed spots, etc., and the occurrence of such inspection materials on crime sites is increasing with the continuous improvement of criminal means. In general, it is difficult to obtain a biological sample with an ideal test result by using a conventional test technique, and one of the difficulties is a contact trace sample. There is currently no hard index that distinguishes trace DNA from traditional STR typing (Gill & Buckleton 2010), but there have been several different discussions of defining it, as defined by the DNA content detected by PCR reactions (polymerase chain reactions), such as < 100pg or < 200pg, as determined by quantitative analysis. This is the case where the number of cells required is 25-50, which is also the case with an extraction efficiency of 100%, so we need to pay attention to samples with a contact trace sample cell number of 20-100.
The existing swab materials for collecting the contact sample are generally as follows: 1. polyester fiber, polyester fiber or artificial silk head, the shank is a swab of plastic or aluminum: it is suitable for collecting virology test specimens. 2. Cotton swab: the kit is suitable for collecting vaginal, cervical and urethral specimens for mycoplasma examination, and is not suitable for collecting bacteria (especially causticized bacteria) and chlamydia examination specimens. 3. Polyester swab and nylon swab: it is suitable for sampling virus and bacteria specimens. 4. Flocking swab: is prepared from nylon fiber by special spraying technology, and is suitable for sampling respiratory viruses and fungi culture specimens. 5. Calcium alginate swab: it is suitable for collecting chlamydia and pertussis Bao Te nasopharyngeal swab, but is not suitable for sampling lipid enveloped virus and cell culture, and is not suitable for sampling neisseria gonorrhoeae.
These different types of swabs described above have less consistent operation. When using a cotton swab, for example, the DNA traces on these contact samples may be transferred to a sterile cotton swab dipped in distilled water, and the trace sites wiped with a wet cotton swab until the traces are transferred to the cotton swab. The so-called "double-swab technique" of wet, dry-swab combination (Sweet at 1997) is one of the most common methods of harvesting cells. The cotton swab is first soaked with sterile distilled water and the surface of the trace is cleaned to hydrate the cells and loosen them from the attached surface. Additional cells were then harvested from the attachment surface with a dry cotton swab. It is generally believed that the rehydrated cells adhere more readily to the dry swab. Unfortunately, this method can limit the amount of DNA recovered due to the poor efficiency of DNA extraction from the cotton swab. A second difficulty in swab collection is that cells can adhere to cotton fibers and are not easily released, and Voorbes 2006 mentions that the need to digest the swab with a cellose to break down cellulose in the cotton can increase DNA recovery. In contrast to conventional swabs, another nylon flocked swab method allows for the recapture of more cells released during extraction and more DNA produced (Benschop et al 2010). The cotton swab of the nylon flocking cotton swab can release more DNA, but has poor water absorption and insufficient adhesiveness to cells, so that the cotton swab can be limited to be used for collecting the contact trace DNA.
In view of the foregoing, there is a need for a high performance swab for forensic contact trace DNA collection that meets the following: (1) The adsorption and collection effects on cells and trace DNA are good; (2) More DNA can be released in the subsequent DNA extraction step; (3) does not affect subsequent PCR amplification.
Disclosure of Invention
Aiming at the defects of the prior art, the invention aims to provide a DNA acquisition swab, and a preparation method and application thereof. The DNA collection swab has the characteristics of porosity, hydrophilicity, cell adhesiveness and easiness in desorption.
To achieve the purpose, the invention adopts the following technical scheme:
in a first aspect, the present invention provides a DNA collection swab, the DNA collection swab comprising a polycaprolactone, a hydrophilic medium, and a carrier;
the hydrophilic medium comprises a saccharide and/or a non-saccharide organic high molecular polymer.
In the invention, polycaprolactone is taken as a framework of a DNA acquisition swab, and a saccharide and/or a non-saccharide organic high molecular polymer is taken as a hydrophilic medium to modify the polycaprolactone framework, and the modified polycaprolactone framework and the hydrophilic medium are composited on a carrier together to form the swab. Polycaprolactone has good biocompatibility, good organic polymer compatibility and good biodegradability, and the hydrophilic medium I is a hydrophilic group for providing polycaprolactone; and a cross-linked network structure is provided between the modified polyurethane and the polycaprolactone, so that the modified polyurethane has strong water absorption property on the polycaprolactone skeleton by the hydrophilic medium, and the formed polyurethane has excellent adsorption and collection effects on cells and trace DNA (the DNA is collected on leather or glass, and the collection effects are excellent), can release more DNA in the subsequent DNA extraction step, does not influence the subsequent PCR amplification (in-vitro DNA amplification technology, is an enzymatic synthesis reaction depending on DNA polymerase in the presence of template DNA, primers and deoxynucleotides), and can be applied to collection of contact trace DNA.
Preferably, the mass ratio of polycaprolactone to hydrophilic medium is 1:10-10:1, and may be, for example, 1:10, 2:10, 3:10, 4:10, 5:10, 6:10, 7:10, 8:10, 9:10, 10:10, 10:9, 10:8, 10:7, 10:6, 10:5, 10:4, 10:3, 10:2, 10:1, etc. Wherein the mass ratio of hydrophilic medium to polycaprolactone can ultimately affect its porosity, hydrophilicity and cell adhesion; if the content is not within the range, the polycaprolactone is too much, the hydrophilicity of the final swab film is insufficient due to regional hydrophobicity, and the formed pore diameter is smaller; if the hydrophilic medium is too much and the polycaprolactone is too little, the rigidity of the whole swab film is reduced, the swab film is in a collapse state in the subsequent alkali treatment process, and a small amount of polycaprolactone cannot form a grid-penetrating state and cannot form a porous composite structure.
Preferably, the weight average molecular weight of the polycaprolactone is 10000-60000, and for example 10000, 20000, 30000, 40000, 50000, 60000, etc. can be used.
Preferably, the saccharide comprises any one or a combination of at least two of a monosaccharide, a disaccharide or a polysaccharide, preferably a polysaccharide. Among them, the saccharide is selected from polysaccharides, because a saccharide with a higher molecular weight can form a porous network with polycaprolactone of a polymer, and monosaccharide has poor effect.
Preferably, the monosaccharides include glucose and/or fructose.
Preferably, the disaccharide comprises trehalose and/or sucrose.
Preferably, the polysaccharide comprises any one or a combination of at least two of chitosan, agarose, dextran, starch, cellulose or carboxymethyl cellulose.
Preferably, the non-saccharide organic high molecular polymer comprises any one or a combination of at least two of polyethyleneimine, polylactic acid, polyvinylpyrrolidone, polystyrene maleic acid ethane sulfonic acid, polyurethane, polyvinyl alcohol, polylactic acid-glycolic acid copolymer, polyprenol, polyether polyol, polyisoprenol, dicyclo alcohol dimer, polycarbonate diol, polycaprolactone diol, acryl polyether or dimer acid polyamide, and is preferably polylactic acid and/or polyvinyl alcohol.
Preferably, the weight average molecular weight of the non-saccharide organic high molecular polymer is 2000-18000, for example, 2000, 4000, 6000, 8000, 10000, 12000, 14000, 16000, 18000, etc. can be mentioned.
Preferably, the weight average molecular weight of the polylactic acid is 14000-16000, for example, 14000, 14200, 14400, 14600, 14800, 15000, 15200, 15400, 15600, 15800, 16000, etc., preferably 15000.
Preferably, the weight average molecular weight of the polyvinyl alcohol is 8000 to 15000, for example, 8000, 9000, 10000, 11000, 12000, 13000, 14000, 15000, etc., preferably 12000.
Preferably, the material of the carrier comprises any one of wood, polyethylene, polypropylene, polytetrafluoroethylene or metal.
Preferably, the metal comprises any one or a combination of at least two of iron, aluminum or an alloy.
Preferably, the alloy comprises any one or a combination of at least two of molybdenum tungsten alloy, niobium tungsten alloy, carbon titanium alloy or carbon tantalum alloy.
Preferably, the carrier is made of iron wires, aluminum wires or alloy materials in a narrow space, and is made of wood or polyethylene, polystyrene or polytetrafluoroethylene materials in a non-narrow space.
Wherein the narrow space refers to a cylindrical die space with a diameter smaller than 5 mm; the non-narrow space refers to a cylindrical mold space having a diameter of 5mm or more.
Preferably, the raw materials for preparing the DNA acquisition swab further comprise an alkali treatment solution. The DNA collecting swab of the invention is treated by alkali treatment liquid, and the DNA collecting swab obtained by the crosslinking reaction is also treated by alkali. Through alkali treatment, ester bonds in the polycaprolactone molecules can be hydrolyzed to form hydrophilic groups carboxyl and hydroxyl, and the carboxyl which is not easy to ionize continuously reacts with alkali to become easily ionized salt, so that the hydrophilic performance of the DNA acquisition swab is further improved.
The pH of the alkali treatment solution is preferably 11 or more, and may be, for example, 11, 11.5, 12, 12.5, 13, 13.5, 14, or the like.
In a second aspect, the present invention provides a method for preparing a DNA collection swab according to the first aspect, the method comprising: and (3) dissolving and mixing polycaprolactone and a hydrophilic medium, and placing the mixture and a carrier in a mould together for molding to obtain the DNA acquisition swab.
Preferably, the preparation method of the DNA acquisition swab comprises the following steps:
(1) Dissolving polycaprolactone in a pre-dissolving agent A to obtain a polycaprolactone solution; dissolving a hydrophilic medium in the pre-dissolving solvent B to obtain a hydrophilic medium solution;
(2) And (3) mixing the polycaprolactone solution obtained in the step (1) with a hydrophilic medium solution, and then placing the mixture and a carrier in a mould together for molding to obtain the DNA acquisition swab.
Preferably, the pre-dissolved solvent a in step (1) comprises any one or a combination of at least two of dichloromethane, chloroform, cyclohexane, toluene, isooctane or glacial acetic acid.
Preferably, the mass volume percentage concentration of the polycaprolactone solution in the step (1) is 1-20%, for example, 1%, 2%, 4%, 6%, 8%, 10%, 12%, 14%, 16%, 18%, 20%, etc. ("mass volume percent concentration" refers to the ratio of the mass of solute in solution to the volume of solvent in solution.)
Preferably, the temperature of the pre-dissolved solvent A in the step (1) is 20 to 90℃and may be, for example, 20℃25℃30℃35℃40℃45℃50℃55℃60℃65℃70℃75℃80℃85℃90 ℃.
Preferably, the pre-dissolving agent A is any one or a combination of at least two of dichloromethane, chloroform or glacial acetic acid. The pre-dissolving solvent A is dichloromethane and/or chloroform, and the polycaprolactone is added and then mechanically stirred at normal temperature; the pre-dissolving solvent A is glacial acetic acid, and the pre-dissolving solvent A needs to be heated and dissolved, wherein the temperature of the heating and dissolving is 60-90 ℃, and the heating time is 1-4h (for example, 1h, 1.5h, 2h, 2.5h, 3h, 3.5h, 4h and the like).
Preferably, the pre-dissolved solvent B in step (1) comprises any one or a combination of at least two of dichloromethane, chloroform, cyclohexane, toluene, isooctane, glacial acetic acid, water, methanol, ethanol, n-propanol, 1, 2-propanediol, glycerol, methyl ether or diethyl ether.
In the invention, the polycaprolactone solution can be stored at 25 ℃ for less than 3 days, and long chains of the polycaprolactone are broken after more than three days, so that an interpenetrating network structure cannot be formed finally.
Preferably, the hydrophilic medium solution in the step (1) has a mass-volume percentage concentration of 2-40%, for example, 2%, 4%, 6%, 8%, 10%, 12%, 14%, 16%, 18%, 20%, 22%, 24%, 26%, 28%, 30%, 32%, 34%, 36%, 38%, 40%, etc. ("mass volume percent concentration" refers to the ratio of the mass of solute in solution to the volume of solvent in solution.)
Preferably, the temperature of the pre-dissolved solvent B in the step (1) is 20 to 90℃and may be, for example, 20℃25℃30℃35℃40℃45℃50℃55℃60℃65℃70℃75℃80℃85℃90 ℃.
Preferably, in the step (2), the carrier is placed in a mold containing the mixed liquor, and the distance between the carrier and the bottom end of the liquid surface of the mixed liquor is more than 0.1cm, for example, the distance between the carrier and the bottom end of the liquid surface of the mixed liquor may be 0.1cm, 0.2cm, 0.3cm, 0.4cm, 0.5cm, etc.
Preferably, the molding temperature in the step (2) is-80 to-20 ℃, and may be, for example, -20 ℃, -30 ℃, 40 ℃, -50 ℃, -60 ℃, -70 ℃, -80 ℃, etc., and the molding time is 0.5 to 2 hours, and may be, for example, 0.5 hours, 0.6 hours, 0.8 hours, 1 hour, 1.2 hours, 1.4 hours, 1.6 hours, 1.8 hours, 2 hours, etc.
Preferably, the DNA collection swab obtained in the step (2) is subjected to freeze drying, cleaning, alkali treatment liquid treatment and sterilization in sequence.
Preferably, the temperature of the freeze-drying treatment is-80 to-20 ℃, for example, -20 ℃, -30 ℃, 40 ℃, -50 ℃, -60 ℃, -70 ℃, -80 ℃ and the like, and the time of the freeze-drying treatment is 1-10h, for example, 1h, 2h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, 10h and the like.
Preferably, the pH of the washed DNA acquisition swab is between 6.8 and 7.2, e.g. 6.8, 6.9, 7, 7.1, 7.2, etc.
The pH of the alkali treatment liquid is preferably 11 or more, and may be, for example, 11, 12, 13, 14, or the like.
Preferably, the sterilization treatment employs ultraviolet sterilization.
Preferably, the preparation method of the DNA acquisition swab comprises the following steps:
(1) Dissolving polycaprolactone in the pre-dissolving agent A to obtain a polycaprolactone solution with the mass-volume percentage concentration of 1-20%; dissolving a hydrophilic medium in a pre-dissolving solvent B to obtain a hydrophilic medium solution with the mass-volume percentage concentration of 2-40%;
(2) Mixing the polycaprolactone solution obtained in the step (1) and the hydrophilic medium solution according to the volume ratio of (1-2): 1, then placing the mixture and the carrier in a mold together, and placing the mold at the temperature of-80 to-20 ℃ for molding for 0.5-2 hours to obtain the DNA acquisition swab;
(3) And (3) freeze-drying the DNA collecting swab obtained in the step (2) at the temperature of minus 80 to minus 20 ℃ for 1 to 10 hours, cleaning until the pH value of the DNA collecting swab is 6.8 to 7.2, soaking the DNA collecting swab in an alkaline treatment solution with the pH value of more than 11 for 12 to 48 hours, cleaning, drying, and then sterilizing under an ultraviolet lamp.
In a third aspect, the present invention provides the use of a DNA collection swab as described in the first aspect for trace DNA detection.
Compared with the prior art, the invention has the following beneficial effects:
(1) The invention takes polycaprolactone as the skeleton of a DNA collecting swab, and uses saccharide and/or non-saccharide organic high molecular polymer as hydrophilic medium to modify the polycaprolactone skeleton, and the polycaprolactone skeleton and the hydrophilic medium are composited on a carrier together to form the swab. The hydrophilic medium modifies the polycaprolactone skeleton and then endows the swab with strong water absorption, so that the swab has excellent effect of adsorbing and collecting cells and trace DNA, can release more DNA in the subsequent DNA extraction step, and does not influence the subsequent PCR amplification;
(2) The water absorption of the DNA acquisition swab can reach more than 180 mu L; in the released cell test: the release rate of 100 cells is up to more than 78%, the release rate of 1000 cells is up to more than 74%, and the release rate of 10000 cells is up to more than 82%.
Drawings
FIG. 1 is a scanning electron microscope image of a DNA acquisition swab prepared in example 1.
FIG. 2 is a scanning electron microscope image of the DNA collection swab prepared in example 2.
FIG. 3 is a short-segment repeat typing plot of a DNA acquisition swab prepared in example 2.
FIG. 4 is a graph showing the comparison of DNA concentration of trace amounts of diluted blood samples extracted from DNA collection swab and cotton swab prepared in example 2.
FIG. 5 is a graph of DNA concentration versus concentration of extracted and desorbed DNA of trace amounts of diluted saliva samples with a DNA collection swab and a cotton swab prepared in example 2.
FIG. 6 is a graph of standard DNA release concentration versus trace amounts of contact samples for the DNA collection swab and the cotton swab prepared in example 2.
FIG. 7 is a graph showing the comparison of DNA concentration of various trace species collected by the DNA collection swab and the cotton swab prepared in example 2.
Detailed Description
The technical scheme of the invention is further described below by the specific embodiments with reference to the accompanying drawings. It should be apparent to those skilled in the art that the examples are merely provided to aid in understanding the present invention and should not be construed as limiting the invention in any way.
Example 1
The embodiment provides a DNA collection swab, and the preparation method of the DNA collection swab comprises the following steps:
(1) Dissolving polycaprolactone with weight average molecular weight of 6 ten thousand in dichloromethane solution at 25 ℃ to prepare polycaprolactone solution with mass and volume percentage concentration of 10%; dissolving chitosan (molecular weight of 20 kD) in glacial acetic acid solution, and heating at 70deg.C for 2 hr to obtain hydrophilic medium solution with mass-volume percentage concentration of 2%;
(2) Uniformly mixing the polycaprolactone solution obtained in the step (1) and the hydrophilic medium solution according to the volume ratio of 1:1 at 25 ℃ for 2 hours to obtain a mixed solution, adding 0.75mL of the mixed solution into an aluminum cylindrical mold (2 mL of liquid can be added into the mold in total), placing an iron wire carrier in the mold containing the mixed solution, placing the iron wire carrier with a space of 0.5cm away from the bottom end of the liquid level, placing the iron wire carrier at 25 ℃ for 30 minutes, and then molding the whole at-60 ℃ for 1 hour to obtain the DNA acquisition swab;
(3) Placing the iron wire carrier connected moulds in a freeze dryer, and freeze drying for 2h at-20 ℃; placing the dried polycaprolactone composite swab in an ethanol water solution with the volume ratio concentration of 50%, cleaning for multiple times until the pH value is 7, and air-drying at 25 ℃; placing the air-dried DNA collection swab in a 2mL centrifuge tube, adding 1mL of 2M sodium bicarbonate (pH 11.5), treating for 30h, repeatedly cleaning with distilled water until the pH is 7, and drying at 50 ℃; and (3) placing the dried DNA collecting swab under an ultraviolet lamp for sterilization treatment for 2 hours, and then placing the DNA collecting swab into a packaging bag without DNase and RNase for preservation.
FIG. 1 is a scanning electron microscope image of the swab surface of example 1, from which it can be seen: the pore size distribution of the surface of the DNA acquisition swab is 20-50 mu m, and the surface of the DNA acquisition swab has more pores, so that the DNA acquisition swab has stronger water absorption and adsorption capacity.
Example 2
The embodiment provides a DNA collection swab, and the preparation method of the DNA collection swab comprises the following steps:
(1) Dissolving polycaprolactone with the weight average molecular weight of 38000 in glacial acetic acid, and heating at 80 ℃ for 2 hours to prepare a polycaprolactone solution with the mass-volume percentage concentration of 15%; dissolving carboxymethyl cellulose with weight average molecular weight of 2 ten thousand in glacial acetic acid solution, and heating and dissolving at 80 ℃ for 2 hours to prepare hydrophilic medium solution with mass and volume percentage concentration of 6%;
(2) Uniformly mixing the polycaprolactone solution obtained in the step (1) and the hydrophilic medium solution according to a volume ratio of 2:1 at 25 ℃ for 2 hours to obtain a mixed solution, adding 0.75mL of the mixed solution into an aluminum cylindrical mold (2 mL of liquid can be added into the mold in total), placing a polytetrafluoroethylene carrier into the mold containing the mixed solution, placing the polytetrafluoroethylene carrier at a space of 0.5cm away from the bottom end of the liquid surface, placing the whole at-20 ℃ for 1 hour after placing the polytetrafluoroethylene carrier at 25 ℃ for 30 minutes, then placing the whole at-30 ℃ for 1 hour, placing the whole at-50 ℃ for 1 hour, and finally forming the mixture at-80 ℃ for 2 hours to obtain the DNA collecting swab;
(3) Placing a polytetrafluoroethylene carrier connected mould in a freeze dryer, and freeze drying for 2 hours at the temperature of minus 20 ℃ to remove redundant solution; placing the dried polycaprolactone composite swab in an ethanol water solution with the volume ratio concentration of 50%, cleaning for multiple times until the pH value is 7, and air-drying at 25 ℃; placing the air-dried DNA collection swab in a 2mL centrifuge tube, adding 1mL of 2M sodium hydroxide, treating for 30h, repeatedly cleaning with distilled water to pH 7, and drying at 50 ℃; and (3) placing the dried DNA collecting swab under an ultraviolet lamp for sterilization treatment for 2 hours, and then placing the DNA collecting swab into a packaging bag without DNase and RNase for preservation.
FIG. 2 is a scanning electron microscope image of the swab surface of example 2, from which it can be seen: the pore size distribution of the surface of the DNA acquisition swab is 5-25 mu m, and the surface of the DNA acquisition swab has more pores, so that the DNA acquisition swab has stronger water absorption and adsorption capacity.
Example 3
The embodiment provides a DNA collection swab, and the preparation method of the DNA collection swab comprises the following steps:
(1) Dissolving polycaprolactone with the weight average molecular weight of 38000 in glacial acetic acid, and heating at 80 ℃ for 2 hours to prepare a polycaprolactone solution with the mass-volume percentage concentration of 15%; dissolving carboxymethyl cellulose with weight average molecular weight of 2 ten thousand in glacial acetic acid solution, and heating and dissolving at 80 ℃ for 2 hours to prepare hydrophilic medium solution with mass and volume percentage concentration of 6%;
(2) Uniformly mixing the polycaprolactone solution obtained in the step (1) and the hydrophilic medium solution according to a volume ratio of 2:1 at 25 ℃ for 2 hours to obtain a mixed solution, adding 0.75mL of the mixed solution into an aluminum cylindrical mold (2 mL of liquid can be added into the mold in total), placing an iron wire carrier in the mold containing the mixed solution, placing the iron wire carrier at a space of 0.5cm away from the bottom end of the liquid surface, placing the iron wire carrier at 25 ℃ for 30 minutes, placing the whole at-20 ℃ for 1 hour, then placing the whole at-30 ℃ for 1 hour, placing the whole at-50 ℃ for 1 hour, and finally forming the iron wire carrier at-80 ℃ for 2 hours to obtain the DNA collecting swab;
(3) Placing the iron wire carrier connected moulds in a freeze dryer, freeze drying for 2 hours at the temperature of minus 20 ℃ and removing redundant solution; placing the dried polycaprolactone composite swab in an ethanol water solution with the volume ratio concentration of 50%, cleaning for multiple times until the pH value is 7, and air-drying at 25 ℃; and (3) placing the air-dried DNA collection swab under an ultraviolet lamp for sterilization treatment for 2 hours, and then filling the DNA collection swab into a packaging bag without DNase and RNase for preservation.
Example 4
This example provides a DNA collection swab differing from example 1 in that the polycaprolactone has a weight average molecular weight of 9000, and otherwise identical to example 1.
Example 5
This example provides a DNA collection swab which differs from example 1 in that the polycaprolactone has a weight average molecular weight of 18 ten thousand, with the other conditions being the same as in example 1.
Example 6
This example provides a DNA collection swab differing from example 1 in that chitosan was replaced with polylactic acid having a weight average molecular weight of 15000, and the other conditions were the same as in example 1.
Example 7
This example provides a DNA acquisition swab differing from example 1 in that chitosan was replaced with polyethyleneimine having a weight average molecular weight of 15000, and the other conditions were the same as in example 1.
Example 8
This example provides a DNA collection swab that differs from example 1 in that chitosan is replaced with trehalose, with the other conditions being the same as example 1.
Example 9
This example provides a DNA collection swab which differs from example 1 in that the polycaprolactone solution and hydrophilic medium solution in step (2) are present in a volume ratio of 10:1, with the other conditions being the same as in example 1.
Example 10
This example provides a DNA collection swab which differs from example 1 in that the polycaprolactone solution and hydrophilic medium solution in step (2) are present in a volume ratio of 1:10, with the other conditions being the same as in example 1.
Example 11
This example provides a DNA collection swab which differs from example 1 in that in step (3) the air-dried DNA collection swab is placed in a 2mL centrifuge tube, 1mL of 0.5M sodium bicarbonate (pH 9.0) is added, and after 30h of treatment, the other conditions are the same as in example 1.
Comparative example 1
The comparative example provides a DNA collection swab, the preparation method of the DNA collection swab comprises the following steps:
(1) Dissolving polycaprolactone with weight average molecular weight of 6 ten thousand in dichloromethane solution at 25 ℃ to prepare polycaprolactone solution with mass and volume percentage concentration of 10%;
(2) Adding 0.75mL of the polycaprolactone solution obtained in the step (1) into an aluminum cylindrical mold (2 mL of liquid can be added into the mold), placing an iron wire carrier into the mold containing the mixed liquid, placing the iron wire carrier at a space of 0.5cm away from the bottom end of the liquid level, placing the iron wire carrier at 25 ℃ for 30min, and then placing the whole at-60 ℃ for molding for 1h to obtain the DNA collecting swab;
(3) Placing the iron wire carrier connected moulds in a freeze dryer, and freeze drying for 2h at-20 ℃; placing the dried polycaprolactone composite swab in an ethanol water solution with the volume ratio concentration of 50%, cleaning for multiple times until the pH value is 7, and air-drying at 25 ℃; placing the air-dried DNA collection swab in a 2mL centrifuge tube, adding 1mL of 2M sodium bicarbonate (pH 11.5), treating for 30h, repeatedly cleaning with distilled water to pH 7, and drying at 50 ℃; and (3) placing the dried DNA collecting swab under an ultraviolet lamp for sterilization treatment for 2 hours, and then placing the DNA collecting swab into a packaging bag without DNase and RNase for preservation.
Comparative example 2
The comparative example provides a DNA collection swab, the preparation method of the DNA collection swab comprises the following steps:
(1) Dissolving polylactic acid with weight average molecular weight of 15000 in glacial acetic acid solution, and heating and dissolving for 2 hours at 70 ℃ to prepare hydrophilic medium solution with mass-volume percentage concentration of 2%;
(2) Adding 0.75mL of the hydrophilic medium solution obtained in the step (1) into an aluminum cylindrical mold (2 mL of liquid can be added into the mold in total), placing an iron wire carrier into the mold containing the mixed liquid, placing the iron wire carrier at a space of 0.5cm away from the bottom end of the liquid level, placing the iron wire carrier at 25 ℃ for 30min, and then placing the whole at-60 ℃ for molding for 1h to obtain the DNA collecting swab;
(3) Placing the iron wire carrier connected moulds in a freeze dryer, and freeze drying for 2h at-20 ℃; placing the dried polycaprolactone composite swab in an ethanol water solution with the volume ratio concentration of 50%, cleaning for multiple times until the pH value is 7, and air-drying at 25 ℃; placing the air-dried DNA collection swab in a 2mL centrifuge tube, adding 1mL of 2M sodium bicarbonate (pH 11.5), treating for 30h, repeatedly cleaning with distilled water to pH 7, and drying at 50 ℃; and (3) placing the dried DNA collecting swab under an ultraviolet lamp for sterilization treatment for 2 hours, and then placing the DNA collecting swab into a packaging bag without DNase and RNase for preservation.
Test example 1
Swab acquisition Water absorbency test
The DNA collection swab prepared in examples 1-11, the DNA collection swab prepared in comparative examples 1-2, and commercial cotton swab, nylon swab, flocked swab, and sponge swab were taken, each swab sample was placed in a 2mL EP tube (microcentrifuge tube) to which 300. Mu.L of deionized water had been added, the swab was allowed to stand for 3min and removed, and the loss of deionized water in the EP tube was weighed to obtain the water absorption of the swab, and the results were recorded in three parallel tests. The specific test results are shown in table 1:
TABLE 1
As can be seen from the test data in Table 1, the DNA collection swab prepared by the method can absorb more than 180 mu L, and the water absorption is stronger than that of a cotton swab, a nylon swab and a flocking swab. Whereas example 3 provides a DNA collection swab which has not been subjected to an alkali treatment, the polymer surface obtained by crosslinking polycaprolactone with a hydrophilic medium has fewer hydrophilic groups and thus has poorer hydrophilic properties, and thus has poorer water absorption properties. In contrast, comparative example 1, which is not compounded with the water absorption of the hydrophilic medium, has no hydrophilic group on the surface of polycaprolactone, and the hydrophobic structure of polycaprolactone itself causes poor water absorption performance of the prepared material.
Test example 2
Swab release cell test
Test sample: the DNA acquisition swab prepared in examples 1 to 11 above, the DNA acquisition swab prepared in comparative examples 1 to 2, and commercial cotton swab, nylon swab, flocked swab, and sponge swab.
The testing method comprises the following steps: preparing a cell sample, wherein the cell sample comprises about 100 cells in each 10 mu L of group A, about 1000 cells in each 10 mu L of group B, about 10000 cells in each 10 mu L of group C, dropwise adding 10 mu L of each group, naturally airing, respectively placing the swabs into a centrifuge tube containing 200 mu L of deionized water, centrifuging and removing the swabs after vortex vibration and uniform mixing, recovering 200 mu L of deionized water, staining the cells by using staining solution, counting under a fluorescence microscope, and judging the release efficiency of the cells. The specific test results are shown in table 2:
TABLE 2
As can be seen from the test data in Table 2, the DNA collecting swab prepared by the invention has a release rate of over 78% for 100 cells, a release rate of over 74% for 1000 cells and a release rate of over 82% for 10000 cells. The invention is illustrated that polycaprolactone is taken as the framework of a DNA collecting swab, and the polycaprolactone framework is modified by taking saccharide and/or non-saccharide organic high molecular polymer as hydrophilic medium, and the polycaprolactone and the hydrophilic medium are composited together on a carrier to form the swab. After the hydrophilic medium modifies the polycaprolactone skeleton, cells can be clung to the surface of the swab, and the cells can be fully and completely infiltrated and cracked by the cracking liquid in the cracking process and can be released to the maximum extent, so that the release efficiency of the cells is remarkably improved, and the adsorption and collection effects of the swab on the cells are formed.
The cotton fibers are mutually entangled to form fiber masses, so that cells are easily wrapped in the fiber masses, and the mutually entangled cotton fibers can prevent effective release of adsorbed cells in the cotton fibers in the cracking process. The test results suggest that the cell release rate of the attached cells from the template DNA on the swab decreases significantly with the increase of the shelf life.
Test example 3
Quantitative results using nucleic acid extraction after swab collection
Test sample: the DNA acquisition swab prepared in examples 1 to 11 above, the DNA acquisition swab prepared in comparative examples 1 to 2, and commercial cotton swab, nylon swab, flocked swab, and sponge swab.
The testing method comprises the following steps: dripping cell sample containing 80-120 cells per 10 μl onto smooth leather, standing, air drying, dipping small amount of PBS solution with each swab, wiping sample point on leather, and applying Qiagen companyDNA Investigator Kit (Cat 952034), in combination with Qiagen EZ1 XL automated sample processor, nucleic acid extraction, adding the swab of the sample into a 2mL centrifuge tube, and adding +.>DNA Investigator Kit 390. Mu.L of lysate and 10. Mu.L of proteinase K solution were lysed for 1h at 56℃at 500rpm, and 150. Mu.L of supernatant was then added to Qiagen EZ1 XL workingThe station was finally eluted with 40 μl deionized water. The eluent was a Quantifiler from Applied Biosystems TM Human DNA Quantification Kit qPCR quantification was performed.
Detection result: after the reaction is finished, the instrument automatically stores the result, the software of the instrument is used for analysis, and the Ct value and the fixed value result of the sample are recorded. The intersection of the amplification curve and the threshold line is called Ct (i.e., cycle threshold, which refers to the number of cycles that the fluorescent signal in the PCR reaction tube undergoes when reaching a set threshold); specific test data are shown in table 3 below (where Ct (first) and Ct (second) refer to values obtained by repeated testing of two parallel samples).
Using 1. Mu.L of the eluate, using Global Filer, applied Biosystems Co TM Express PCR Amplification Kit STR amplification was performed using 3500XL electrophoresis apparatus from Applied Biosystems company, geneMapper software analysis, FIG. 3 is a short segment repeat typing chart of the DNA collection swab prepared in example 2, a bit STR peak type data chart, the number under each locus representing the number of times of its repeat, it can be seen from the chart that STR has good peak integrity, each peak height exceeding 1000, demonstrated high DNA collection, and more DNA was released in the DNA extraction step without affecting the subsequent PCR amplification.
TABLE 3 Table 3
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As can be seen from the test data in Table 3, the DNA collection swab prepared by the invention has relatively smaller extraction efficiency of DNA from Ct quantitative value than the swab prepared by comparative examples 1-2 and the Ct value of each commercial swab, and has relatively better cleavage effect because the swab prepared by comparative examples 1-2 and each commercial swab have low release rate of DNA process when collecting samples, resulting in smaller number of finally obtained nucleic acid templates. The swab prepared by the method disclosed by the invention takes polycaprolactone as a framework of a DNA acquisition swab, and uses the saccharide and/or non-saccharide organic high-molecular polymer as a hydrophilic medium to modify the polycaprolactone framework, so that the amount of nucleic acid released by the swab formed by compounding the polycaprolactone on a carrier is more, the cracking effect is relatively good, and the extraction efficiency is high.
Test example 4
Investigation of acquisition and desorption efficiency
Test sample: the DNA collection swab and the cotton swab prepared in example 2 above.
The testing method comprises the following steps:
(1) Investigation of collection and desorption efficiency of trace diluted blood samples: after the conventional sampled blood sample is diluted by 100 times, 10 mu L of the diluted blood sample is dripped on smooth leather, the mixture is stood and air-dried, a small amount of PBS solution is dipped by using each swab, and then a sample point on the leather is wiped, wherein the used extraction reagent, instrument and quantitative reagent are consistent with those of test example 3;
(2) Collection and desorption efficiency investigation of trace diluted saliva samples: diluting a conventional saliva sample by 50 times, dripping 10 mu L of the saliva sample onto smooth leather, standing and air-drying, dipping a small amount of PBS solution by using each swab, and wiping a sample point on the leather, wherein the used extraction reagent, instrument and quantitative reagent are consistent with those of test example 3;
(3) Investigation of Standard DNA Release efficiency: DNA release efficiency was evaluated using standard DNA9947A, 20. Mu.L of 10 ng/. Mu.L of standard DNA was added to the swab head, and after natural air drying, 200. Mu.L of deionized water was added and treated at 1500rpm at 56℃for 25 minutes, and the DNA in deionized water was analyzed.
The specific test results are shown in table 4:
TABLE 4 Table 4
As shown in the test results of Table 4, the extraction efficiency of the DNA collection swab prepared by the invention on trace diluted blood samples is more than 80%, and the desorption concentration is more than 0.048 ng/. Mu.L; the extraction efficiency of the trace diluted saliva sample is above 89%, and the desorption concentration is above 0.136 ng/. Mu.L; the release efficiency of the DNA sample is over 90 percent, and the DNA concentration is over 0.09 ng/. Mu.L. The DNA collection swab prepared by the method can adsorb cells to a greater extent when a sample is collected, trace components in the sample can be efficiently released when the sample is subjected to cracking digestion, and the trace components are not retained in the swab. Wherein, fig. 4 is a graph of comparison of DNA concentration of trace amounts of diluted blood samples extracted and desorbed from the DNA collection swab and the cotton swab prepared in example 2. FIG. 5 is a graph of DNA concentration versus concentration of extracted and desorbed DNA of trace amounts of diluted saliva samples with a DNA collection swab and a cotton swab prepared in example 2. FIG. 6 is a comparison of standard DNA release concentration of the DNA collection swab and the cotton swab prepared in example 2 to trace contact samples, and more clearly shows that trace components in the samples are easy to stay in entangled fiber clusters, and the DNA extraction effect is affected.
Test example 5
Investigation of trace contact sample collection efficiency
Test sample: the DNA acquisition swab prepared in examples 1 to 11 above, the DNA acquisition swab prepared in comparative examples 1 to 2, and commercial cotton swab, nylon swab, flocked swab, and sponge swab.
The testing method comprises the following steps: the above swabs were used to collect and evaluate trace amounts of contact material, several cell phones were selected for wiping, and specific test results are shown in table 5 (wherein sample one refers to volunteer one cell phone, sample two refers to volunteer two cell phone, and sample three refers to volunteer three cell phone):
TABLE 5
As shown in the test results of Table 5, the DNA collecting swab prepared by the invention has excellent collecting effect on cells and trace DNA, the collecting concentration of the sample I is more than 0.008 ng/. Mu.L, the collecting concentration of the sample II is more than 0.03 ng/. Mu.L, and the collecting concentration of the sample III is more than 0.023 ng/. Mu.L. In order to more intuitively show that the DNA collection swab prepared by the present invention has excellent effect of collecting cells and trace amounts of DNA, fig. 7 is a graph showing comparison of DNA concentration collected by different trace amounts of substances of the DNA collection swab prepared in example 2, and from fig. 7, it is more obvious that the average concentration of DNA collected by the DNA collection swab prepared in the present invention is much higher than that of the cotton swab for the first sample and the second sample, and the efficiency of the DNA collection swab prepared in example 2 is still 20.3% higher than that of the cotton swab for the third sample. The invention is characterized in that polycaprolactone is taken as a framework of a DNA collecting swab, and saccharide and/or non-saccharide organic high molecular polymer is taken as a hydrophilic medium to modify the polycaprolactone framework, and the polycaprolactone framework and the hydrophilic medium are composited on a carrier together to form the swab, so that the collecting effect of the swab on cells and trace DNA is excellent.
The applicant states that the present invention is illustrated by the above examples of DNA collection swabs, methods of making and using the same, but the present invention is not limited to, i.e. it is not meant that the present invention must be practiced in dependence upon, the above examples. It should be apparent to those skilled in the art that any modification of the present invention, equivalent substitution of raw materials for the product of the present invention, addition of auxiliary components, selection of specific modes, etc., falls within the scope of the present invention and the scope of disclosure.
Claims (11)
1. The DNA acquisition swab is characterized in that the preparation raw materials of the DNA acquisition swab comprise polycaprolactone, a hydrophilic medium and a carrier;
the hydrophilic medium is polysaccharide;
the mass ratio of the polycaprolactone to the hydrophilic medium is 2:10-10:2;
the weight average molecular weight of the polycaprolactone is 10000-60000;
the preparation raw materials of the DNA acquisition swab further comprise alkali treatment liquid;
the DNA acquisition swab is prepared by the following method: and (3) dissolving and mixing polycaprolactone and a hydrophilic medium, and placing the mixture and a carrier in a mould together for molding to obtain the DNA acquisition swab.
2. The DNA collection swab of claim 1, wherein the polysaccharide comprises any one or a combination of at least two of chitosan, agarose, dextran, starch, cellulose, or carboxymethyl cellulose.
3. The DNA collection swab of claim 1, wherein the carrier comprises any one of wood, polyethylene, polypropylene, polytetrafluoroethylene, or metal.
4. The DNA collection swab of claim 3, wherein the metal comprises any one or a combination of at least two of iron, aluminum, or an alloy.
5. The DNA collection swab of claim 4, wherein the alloy comprises any one or a combination of at least two of a molybdenum tungsten alloy, a niobium tungsten alloy, a carbon titanium alloy, or a carbon tantalum alloy.
6. The DNA collection swab of claim 1, wherein the pH of the alkaline treatment solution is above 11.
7. The method of preparing a DNA collection swab according to any one of claims 1 to 6, wherein the method of preparing a DNA collection swab comprises the steps of:
(1) Dissolving polycaprolactone in a pre-dissolving agent A to obtain a polycaprolactone solution; dissolving a hydrophilic medium in the pre-dissolving solvent B to obtain a hydrophilic medium solution;
(2) And (3) mixing the polycaprolactone solution obtained in the step (1) with a hydrophilic medium solution, and then placing the mixture and a carrier in a mould together for molding to obtain the DNA acquisition swab.
8. The method of preparing a DNA collection swab of claim 7, wherein the pre-solubilising agent a in step (1) comprises any one or a combination of at least two of dichloromethane, chloroform, cyclohexane, toluene, isooctane or glacial acetic acid;
the mass volume percentage concentration of the polycaprolactone solution in the step (1) is 1-20%;
the temperature of the pre-dissolved solvent A dissolved in the step (1) is 20-90 ℃;
the pre-dissolved solvent B in the step (1) comprises any one or a combination of at least two of dichloromethane, chloroform, cyclohexane, toluene, isooctane, glacial acetic acid, water, methanol, ethanol, n-propanol, 1, 2-propanediol, glycerol, methyl ether or diethyl ether;
the mass volume percentage concentration of the hydrophilic medium solution in the step (1) is 2-40%;
the temperature of the pre-dissolved solvent B dissolved in the step (1) is 20-90 ℃.
9. The method according to claim 7, wherein the molding temperature in the step (2) is-80 to-20 ℃, and the molding time is 0.5 to 2 hours;
the DNA collecting swab obtained in the step (2) is sequentially subjected to freeze drying, cleaning, alkali treatment liquid treatment and sterilization treatment;
the temperature of the freeze drying treatment is-80 to-20 ℃, and the time of the freeze drying treatment is 1-10h;
the pH value of the cleaned DNA collection swab is 6.8-7.2;
the pH value of the alkali treatment liquid is above 11;
the sterilization treatment adopts ultraviolet sterilization.
10. The method of preparing a DNA collection swab as recited in claim 7, wherein the method of preparing a DNA collection swab comprises the steps of:
(1) Dissolving polycaprolactone in the pre-dissolving agent A to obtain a polycaprolactone solution with the mass-volume percentage concentration of 1-20%; dissolving a hydrophilic medium in a pre-dissolving solvent B to obtain a hydrophilic medium solution with the mass-volume percentage concentration of 2-40%;
(2) Mixing the polycaprolactone solution obtained in the step (1) and the hydrophilic medium solution according to the volume ratio of (1-2): 1, then placing the mixture and the carrier in a mold together, and placing the mold at the temperature of-80 to-20 ℃ for molding for 0.5-2 hours to obtain the DNA acquisition swab;
(3) Freeze-drying the DNA collecting swab obtained in the step (2) at-80 to-20 ℃ for 1-10 hours, cleaning until the pH value of the DNA collecting swab is 6.8-7.2, soaking in an alkaline treatment solution with the pH value of more than 11 for 12-48 hours, cleaning, drying, and then sterilizing under an ultraviolet lamp;
the pre-dissolved solvent A in the step (1) comprises any one or a combination of at least two of dichloromethane, chloroform, cyclohexane, toluene, isooctane and glacial acetic acid;
the pre-dissolving agent B in the step (1) comprises any one or a combination of at least two of methylene dichloride, chloroform, cyclohexane, toluene, isooctane, glacial acetic acid, water, methanol, ethanol, n-propanol, 1, 2-propylene glycol, glycerol, methyl ether or diethyl ether.
11. Use of a DNA collection swab according to any one of claims 1-6 in trace DNA detection.
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